Nitrogen retention disorders associated with elevated ammonia levels include urea cycle disorders (UCDs), hepatic encephalopathy (HE), and advanced kidney disease or kidney failure, often referred to as end-stage renal disease (ESRD).
UCDs include several inherited deficiencies of enzymes or transporters necessary for the synthesis of urea from ammonia, including enzymes involved in the urea cycle. The urea cycle is depicted in FIG. 1, which also illustrates how certain ammonia-scavenging drugs act to assist in elimination of excessive ammonia. With reference to FIG. 1, N-acetyl glutamine synthetase (NAGS)-derived N-acetylglutamate binds to carbamyl phosphate synthetase (CPS), which activates CPS and results in the conversion of ammonia and bicarbonate to carbamyl phosphate. In turn, carbamyl phosphate reacts with ornithine to produce citrulline in a reaction mediated by ornithine transcarbamylase (OTC). A second molecule of waste nitrogen is incorporated into the urea cycle in the next reaction, mediated by arginosuccinate synthetase (ASS), in which citrulline is condensed with aspartic acid to form argininosuccinic acid. Argininosuccinic acid is cleaved by argininosuccinic lyase (ASL) to produce arginine and fumarate. In the final reaction of the urea cycle, arginase (ARG) cleaves arginine to produce ornithine and urea. Of the two atoms of nitrogen incorporated into urea, one originates from free ammonia (NH4+) and the other from aspartate. UCD individuals born with no meaningful residual urea synthetic capacity typically present in the first few days of life (neonatal presentation). Individuals with residual function typically present later in childhood or even in adulthood, and symptoms may be precipitated by increased dietary protein or physiological stress (e.g., intercurrent illness). For UCD patients, lowering blood ammonia is the cornerstone of treatment.
HE refers to a spectrum of neurologic signs and symptoms believed to result from hyperammonemia, which frequently occur in subjects with cirrhosis or certain other types of liver disease. HE is a common manifestation of clinically decompensated liver disease and most commonly results from liver cirrhosis with diverse etiologies that include excessive alcohol use, hepatitis B or C virus infection, autoimmune liver disease, or chronic cholestatic disorders such as primary biliary cirrhosis. Patients with HE typically show altered mental status ranging from subtle changes to coma, features similar to patients with UCDs. It is believed that an increase in blood ammonia due to dysfunctional liver in detoxifying dietary protein is the main pathophysiology associated with HE (Ong 2003).
ESRD results from a variety of causes including diabetes, hypertension, and hereditary disorders. ESRD is manifested by accumulation in the bloodstream of substances normally excreted in the urine, including but not limited to urea and creatinine. This accumulation in the bloodstream of substances, including toxins, normally excreted in the urine is generally believed to result in the clinical manifestations of ESRD, sometimes referred to also as uremia or uremic syndrome. ESRD is ordinarily treated by dialysis or kidney transplantation. To the extent that urea, per se, contributes to these manifestations and that administration of a phenylacetic (PAA) prodrug may decrease synthesis of urea (see, e.g., Brusilow 1993) and hence lower blood urea concentration, PAA prodrug administration may be beneficial for patients with ESRD.
Subjects with nitrogen retention disorders whose ammonia levels and/or symptoms are not adequately controlled by dietary restriction of protein and/or dietary supplements are generally treated with nitrogen scavenging agents such as sodium phenylbutyrate (NaPBA, approved in the United States as BUPHENYL® and in Europe as AMMONAPS®), sodium benzoate, or a combination of sodium phenylacetate and sodium benzoate (AMMONUL®). These are often referred to as alternate pathway drugs because they provide the body with an alternate pathway to urea for excretion of waste nitrogen (Brusilow 1980; Brusilow 1991). NaPBA is a PAA prodrug. Another nitrogen scavenging drug currently in development for the treatment of nitrogen retention disorders is glyceryl tri-[4-phenylbutyrate] (HPN-100), which is described in U.S. Pat. No. 5,968,979. HPN-100, which is commonly referred to as GT4P or glycerol PBA, is a prodrug of PBA and a pre-prodrug of PAA. The difference between HPN-100 and NaPBA with respect to metabolism is that HPN-100 is a triglyceride and requires digestion, presumably by pancreatic lipases, to release PBA (McGuire 2010), while NaPBA is a salt and is readily hydrolyzed after absorption to release PBA.
HPN-100 and NaPBA share the same general mechanism of action: PBA is converted to PAA via beta oxidation, and PAA is conjugated enzymatically with glutamine to form phenylacetylglutamine (PAGN), which is excreted in the urine. The structures of PBA, PAA, and PAGN are set forth below:

The clinical benefit of NaPBA and HPN-100 with regard to nitrogen retention disorders derives from the ability of PAGN to effectively replace urea as a vehicle for waste nitrogen excretion and/or to reduce the need for urea synthesis (Brusilow 1991; Brusilow 1993). Because each glutamine contains two molecules of nitrogen, the body rids itself of two waste nitrogen atoms for every molecule of PAGN excreted in the urine. Therefore, two equivalents of nitrogen are removed for each mole of PAA converted to PAGN. PAGN represents the predominant terminal metabolite, and one that is stoichiometrically related to waste nitrogen removal, a measure of efficacy in the case of nitrogen retention states.
In addition to nitrogen retention states, PAA prodrugs may be beneficial in a variety of other disorders for which PBA and/or PAA are believed to modify gene expression and/or exert post-translational effects on protein function. In the case of maple syrup urine disease (MSUD, also known as branched-chain ketoaciduria), for example, the apparently beneficial effect of NaPBA in lowering plasma levels of branched chain amino acids is reported to be mediated by PBA-induced inhibition of the kinase that regulates activity of branched chain alpha-keto acid dehydrogenase complex or BCKDC. BCKDC is the enzyme that normally breaks down branched-chain amino acids and is genetically defective in MSUD patients (Bruneti-Pieri 2011). Similarly, the putative beneficial effects of PAA prodrugs for the treatment of cancer (Chung 2000), neurodegenerative diseases (Ryu 2005), and sickle cell disease (Perrine 2008) all involve alteration of gene expression and/or post-translational effects on protein function via PBA and/or PAA.
Numerous publications reports adverse events following administration of PBA and/or PAA (Mokhtarani 2012), and PAA is reported to cause reversible toxicity when present in high levels in circulation. While many of these publications have not recorded PAA blood levels and/or temporally correlated adverse events with PAA levels, toxicities such as nausea, headache, emesis, fatigue, weakness, lethargy, somnolence, dizziness, slurred speech, memory loss, confusion, and disorientation have been shown to be temporally associated with PAA levels ranging from 499-1285 μg/mL in cancer patients receiving PAA intravenously, and these toxicities have been shown to resolve with discontinuation of PAA administration (Thiebault 1994; Thiebault 1995). Therefore, when administering PAA prodrugs for treatment of nitrogen retention disorders and other conditions, it is important to optimize dosing so as to achieve the desired therapeutic effect while minimizing the risk of PAA associated toxicity.